Project Summary/Abstract The lymphatic system, with its intrinsic physiologic functions in cellular homeostasis, immune cell trafficking, and lipid absorption, contributes significantly to the pathogenesis of metabolic disorders. Lymphatic dysfunction shows strong positive correlation with obesity and results in increased susceptibility to lymphedema. Defective lymphatic vessels by lacking one allele of Prox1, a transcription factor that regulates lymphatic endothelial cell differentiation, induces adult-onset obesity, suggesting that lymphatic dysfunction directly contributes to the development of metabolic disorders. In contrast, metabolic diseases exhibit lymphatic dysregulation that contributes to the development of further pathological conditions including inflammation, impaired lipid absorption, or elevated chyle accumulation coupled with increased susceptibility of edema. However, how lymphatic dysfunctions aggravate metabolic diseases, and conversely, how metabolic disorders induce lymphatic dysfunctions are unknown but highly relevant medical questions. Our preliminary data show that deficiency of Foxc2, a critical regulator of lymphangiogenesis, elevates VEGFR3 signaling and causes an increase in lymphangiogenesis both in vivo and in vitro. In addition, loss of lymphatic endothelial Foxc2 enhanced collecting lymphatic function and lymph flow in adult mice. Hence, we will identify 1) the therapeutic potential of targeting Foxc2 in lymphatic dysfunction in metabolic disorders and 2) the underlying molecular mechanisms by which Foxc2 regulates adult lymphangiogenesis and lymphatic function. Using the innovative conditional lymphatic endothelial specific Foxc2 gain-of-function or Foxc2 loss-of-function mice, we will test the direct role of Foxc2 in pathophysiological lymphatic regeneration and function in metabolic diseases. In addition, we will utilize highly sensitive NIR imaging approaches to assess in vivo lymph transport in our live transgenic animals. Finally, we will determine underlying molecular mechanisms by which Foxc2 regulates lymphangiogenesis and lymphatic function. We will use Chip, immunoblotting, co-immunoprecipitation, and luciferase reporters to complete this in- depth investigation. Thus, this proposal will identify important targets that modulate lymphangiogenesis and lymphatic function in mature lymphatic vessels, which could potentially provide diagnostic tools or therapies to improve lymph transport and, consequently mitigate lymphatic dysfunctions in patients with Metabolic Syndrome or secondary lymphedema.